Jundishapur Journal of Microbiology
Volume:16 Issue: 2, Feb 2023

Brucellosis is a zoonotic disease with different clinical symptoms. Its early diagnosis is essential to prevent severe complications. Due to the limitations of serological diagnostic methods, the polymerase chain reaction (PCR) method has become important in the diagnosis of the disease.

Our study aimed to evaluate the PCR method in patients with suspected brucellosis and compare it with serological tests.

This cross-sectional study was performed on 90 febrile patients with clinical features of brucellosis who were examined by an infectious disease specialist. A total of 90 serum samples were collected from the suspected brucellosis patients admitted to the hospital and were analyzed by serological (Rose Bengal) and PCR tests. Then, each method’s results were recorded and compared with each other.

According to serological test results, 45 samples were negative, and 45 were positive. Then, among the serology-positive patients, all had positive PCR results. However, 40 out of 45 patients had a positive PCR test in serology-negative patients. According to this study, the sensitivity of PCR in diagnosing human brucellosis with the serology-positive test is 100%, and with the negative serology test is 88.9%. Therefore, the sensitivity of PCR is higher than that of serology tests in patients, which was 50% in this study.

The PCR test can be a valuable diagnostic method for patients with negative serologic test results.

Community-acquired urinary tract infection is among the most common infections in older adults. Regardless of age, the most frequently detected causative microorganism is Escherichia coli. In parallel with the increase in antibiotic use, the frequency of community-acquired extended-spectrum beta-lactamase-producing E. coli (ESBL-E. coli) has reached critical levels. The use of empirical antibiotic therapy is determined by assessing patient-based risk factors. Therefore, knowing the risk factors and the frequency of antimicrobial resistance can guide the treatment to shape the treatment.

This studyaimedto determine the risksandresistance frequencies to guide the empirical treatment selection for ESBL-E. coli-associated urinary tract infection (UTI) in elderly patients.

This study is a retrospective cohort study. It was carried out between 2011 - 2019. Escherichia coli growth of  105 colonyforming units (cfu)/mL in urine culture was included in 815 patients aged 65 and over who applied to outpatient clinics.

Two hundred and sixty (31.9%) of the patients had ESBL-E. coli. In ESBL-E. coli, antimicrobial resistance rates were highest (100%) for penicillins + -lactamase inhibitors. The lowest resistance rates were determined for carbapenems, aminoglycosides, phosphonic acid, and nitrofurantoins. Risk factors for ESBL-producing bacteria were determined. These were the presence of benign prostatic hypertrophy, antibiotic use in the last three months, history of UTI in the last year, urinary catheter uses in the last year, male gender, and hospitalization in the last year (P < 0.05). The only independent risk factor was a history of UTI in the last year, which increased the risk of ESBL by 2.8 times.

Carbapenems can be chosen as parenteral options, and phosphonic acids and nitrofurantoin as oral options for empirical antibiotic treatment, especially in patients with a history of UTI in the past year.

Since common drug therapies cannot eradicate Candida biofilm, extensive studies are required to develop more effective antifungal compounds and identify their mechanism of action against Candida biofilm. Peganum harmala L. is a traditional medicinal plant, the seeds of which have been used to treat various diseases.

This study aimed to investigate the anti-biofilm mechanisms of P. harmala extract (PHE) and the expression of CAT1, EFG1, and BCR1 genes involved in oxidative stress response and biofilm formation in Candida albicans.

Anti-biofilm activity of PHE was evaluated by crystal violet assay to determine biofilm formation on 33 C. albicans isolates. Finally, a real-time polymerase chain reaction was performed to analyze the effect of PHE on the expression of CAT1, EFG1, and BCR1 genes in C. albicans.

This study determined the minimum biofilm eradication concentration (MBEC) of 15 isolates in concentrations between 0.49 - 3.9g/mL of P. harmala extract. Statistical analysis showed that the exposure of C. albicans biofilm to PHE significantly reduced the expression of CAT1 mRNA in C. albicans isolates (P = 0.0068). However, no significant difference was observed in the expression of EFG1 and BCR1 genes.

The results demonstrated that PHE significantly decreased CAT1 expression in C. albicans cells treated with the herbal extract. PHE is likely to accumulate hydrogen peroxide (H2O2) by reducing CAT1 expression and disrupting the prooxidant/ antioxidant balance that leads to the overproduction of reactive oxygen species (ROS) and can cause damage to cellular components and eventually destroy C. albicans biofilm.

The treatment of Staphylococcus aureus infections has become a public health crisis due to the extensive development of antimicrobial resistance. Antimicrobial peptides (AMPs) have been introduced as promising naturally-derived antimicrobial alternatives to antibiotics. LL-37 and oncorhyncin II are 2 AMPs with notable proven antibacterial effects.

This study aimed to produce recombinant LL-37 and oncorhyncin II and investigate their synergistic effects on S. aureus (ATCC25923).

The synthetic genes of LL-37 and oncorhyncin II were individually ligated into the pET32a expression vector. Transformed pET32a was introduced into Escherichia coli BL21 as an expression host. The protein expression and purification steps were optimized, and the biological effectiveness of the peptides was evaluated by assessing the minimum inhibitory concentration (MIC), time-kill, and growth kinetic tests against S. aureus.

The MIC assay confirmed the effective antibacterial performances of LL-37 and oncorhyncin II against S. aureus at 30.6 and 47.93 g/mL, respectively. The peptides’ synergistic activity was validated by the checkerboard method. A combination of LL-37 and oncorhyncin II at 2  MIC showed a sharp decline of the viable cells with over 3-time reductions in log 10 colony-forming units (CFU)/mL within the first 5 hours. The growth kinetic results confirmed the high effectiveness of the peptides’ combination in eliminating the bacterial inoculum turbidity by 50% reduction during the first hour of exposure.

The produced recombinant LL-37 and oncorhyncin II showed effective antimicrobial function against S. aureus. The synergistic performance of the peptides was repeatedly confirmed through checkerboard, time-kill, and growth kinetic assays.

Crimean-Congo hemorrhagic fever virus (CCHFV) is a highly lethal virus that causes hemorrhagic fever inhumansand is endemic inmanycountries, including Iran. Therefore, fast, accurate, and reliable diagnosis is crucial for patient management and outbreak control.

This study aims to optimize a TaqMan multiplex real-time RT-PCR for the rapid and specific diagnosis of CCHFV.

In this study, the L (NC_005301.3) and S (NC_005302.1) fragments were used as reference sequences for blast analysis. The L and S sequence segments of CCHFV with more than 90% identity from different areas were downloaded from the Genbank database. Primers and probes were designed based on the best-conserved regions of CCHFV L and S sequence segments. To construct the plasmid, a 1751 bp fragment from the MS2 phage that was previously amplified using cloning primers was inserted into the pET-32a plasmid. The S and L segments of the CCHFV, which were 110 bp and 135 bp, respectively, were inserted downstream of the MS2 phage sequence from HindIII to NotI. The Viral-like particles (VLPs) were produced in Escherichia coli, strain BL-21(DE3), in the presence of 1 mM Isopropyl -D-1-thiogalactopyranoside (IPTG). The stability of VLP particles was confirmed in the presence of the ribonuclease enzyme. The fabrication of VLPs was approved by transmission electron microscopy (TEM) with negative staining (1% phosphotungstic acid). To validate the specificity of the primers and probes sequences, we compared them to the NCBI database and tested them experimentally using extracted DNA and RNA samples from healthy subjects and an infectious panel.

The VLPs showed complete resistance in the presence of the ribonuclease enzyme, and the TEM results confirmed that the VLPs were correctly produced. The TaqMan multiplex real-time RT-PCR confirmed that the primers and probes were designed correctly and were completely specific to the CCHFV. The limit of detection (LOD) of the multiplex assay for the L and S genes was one copy of the VLPs per L.

This TaqMan assay is reliable for amplifying CCHFV due to its design on conserved regions of the CCHFV sequences, which have minimal variability and high specificity.

The evidence has shown the relationship between the microbiota of the face and several skin conditions. However, for rosacea patients, the changes in the facial skin microbiota still remain unknown.

This study was performed to explore the correlation between the facial skin microbiota and rosacea and analyze and characterize the facial skin microbiota of rosacea patients in comparison to healthy controls using 16S rDNA amplicon sequencing.

A total of 27 rosacea patients and 25 healthy controls were matched. TheDNAwas extracted from participants’ skin swabs taken from the nose, chin, forehead, and bilateral cheeks. The V3V4 region of the 16S rRNA gene was sequenced using Illumina MiSeq technology. The diversity of the face skin microbiota was examined using alpha and beta diversity. Utilizing linear discriminant analysis effect size (LEfSe), the quantitative study of biomarkers in the two groups was carried out. Clusters of orthologous groups and Kyoto encyclopedia of genes and genomes function predictions were made at the genus level utilizing phylogenetic investigation of communities by reconstruction of unobserved states.

The alpha diversity of the facial skin microbiota increased significantly in rosacea patients, and beta diversity showed substantial differences between the rosacea and healthy control groups. The facial skin microbiota community structure changed in rosacea patients; however, the dominant strains were the same as in healthy controls, both being Propionibacterium acnes and Staphylococcus epidermidis. The LEfSe demonstrated that Xanthomonas, Acinetobacter, and Pseudomonas were enriched in the rosacea patients; nevertheless, Corynebacterium, Finegoldia, and Peptoniphilus were enriched in the healthy controls. The rosacea patients showed significantly decreased expression in the pathways of membrane transport, carbohydrate metabolism, metabolic diseases, amino acid transport and metabolism, carbohydrate transport and metabolism, transcription, and inorganic ion transport and metabolism.

The facial skin microbiota diversity and community structure changed, and the expression of several metabolic pathways was downregulated in the rosacea patients in comparison to the healthy controls, which might outline new strategic methods for the surveillance, diagnosis, and treatment of rosacea.